Sensorineural hearing loss as a result of ototoxic agents, noise exposure, and aging affects millions of people in the United States alone. Considerable research is underway that seeks to protect inner ear sensory hair cells from these challenges. Much of this work is based on our rapidly evolving understanding of cell death and cell survival pathways in other organs and cell-based model systems. It has been shown that ototoxic cell death in the inner ear involves reactive oxygen species and can be attenuated using anti- oxidants. Apoptotic mediators such as caspases have been shown to be involved. However, there is limited knowledge of upstream pathways of ototoxic hair cell death or of the relationship between reactive oxygen species and downstream cell death pathways. The signal transduction and activation of transcription (STAT) proteins are now known to be important mediators of cell death. Multiple studies have shown a direct link between reactive oxygen species and cell death involving STATs in a variety of cell types. STAT1 in particular has been shown to promote cell death in most cell types. However, the possible role of STAT1 in death of mechanosensory hair cells has not yet been elucidated. Experiments proposed here will investigate the role of STAT1 in death of inner ear sensory hair cells upon exposure to the ototoxic drug cisplatin. Preliminary data demonstrate that STAT1 is activated in mouse utricular hair cells following exposure to toxic doses of cisplatin and that an inhibitor of STAT1 activation can reduce cisplatin-induced hair cell death in this preparation. Experiments described in this proposal will further characterize this activation and will determine whether absence of STAT1, via pharmacologic inhibition or genetic knockout, provides protection of hair cells from cisplatin toxicity. Utricular cultures from mature mice will be exposed to cisplatin with or without an inhibitor of STAT1 for different time periods, followed by immunohistochemistry and western blot to investigate the extent of STAT1 activation. Additional utricular explants will be cultured with cisplatin with or without the STAT1 inhibitor for 24 hours followed by hair cell counts to determine whether these inhibitors prevent or reduce hair cell death. Next, utricular explants from wildtype and STAT 1-knockout mice will be exposed to cisplatin to determine whether genetic absence of STAT1 confers hair cell protection. These studies will help elucidate the role of an important cell death pathway in hearing loss as a result of toxic medications. This will add to our current understanding of hair cell death mechanisms, which is necessary for the identification of therapeutic targets for the prevention of hearing loss. [unreadable] [unreadable] [unreadable] [unreadable]